A. Dynorphin A
Dynorphin was discovered in 1979 as being an endogenously potent opioid peptide (PNAS, USA 76:6666–6670: 1979). The pharmacological actions of dynorphins are quite vast. Dynorphin A and related peptides have been shown to be moderately effective in non-thermal and mechanical analgesia when administered ICV (Dubner, R., Trends Neurosci. 15:96–103, 1992) and have also been used in the clinic for the treatment of intractable pain in cancer patients (Wen, H. L., Central and peripheral endorphins: basic and clinical aspects, New York: Raven Press; 1993:319–323). Dynorphins are also known to have an effect on the cardiovascular system via the central and peripheral nervous systems (Dumont, M. 37:1–33, 1996). Moreover, Dynorphin A has an immunomodulatory activity. When administered to mice, Dynorphin A enhanced phagocytosis in the mouse peritoneal macrophages. This phagocytic activity was not inhibited by naloxone treatment suggesting the involvement of a non-opioid receptor (J. Neuroimmunol. 60:37–43, 1995).
Even though, Dynorphin A was first described as a potent opioid peptide with selectivity for kappa opioid receptor, some of its pathological and physiological actions have been proposed to be mediated by non-opioid receptors. Hence, here we described for the first time that Dynorphin A is able to activate and bind to the MAS receptor.
The human MAS oncogene receptor was first isolated in 1986 via a tumorgenicity assay in nude mice (Young, M., Cell, 45:711–719, 1986). This receptor codes for a seven-transmembrane domain protein and within its coding sequence a hallmark feature is present such as the NPY motif in the seventh transmembrane domain. The MAS receptor is expressed in the ventral nervous system (CNS) with highest signal of the mRNA observed in the hippocampus, cortex, cerebellum, piriform cortex and olfactory bulb. The mRNA for this receptor has also been detected in peripheral tissues including the testis, kidney, and heart (FEBS Letters, 357, 27–32, 1995). The expression of MAS receptor is highly regulated during development and neuronal activity. Interestingly, mice lacking the MAS protooncogene not only displayed an increased anxiety but long-term potentiation was prolonged without affecting the gross morphology of the hippocampus (JBC, 273,11867–11873, 1998).
Hence, drugs targeted at the MAS receptor as agonists, antagonists or inverse agonists could be potentially used for treating problems of long-term memory neuropathic and inflammatory disorders, as well as cardiovascular dysfunction.
B. G Protein-Coupled Receptors
G protein coupled receptors (GPCRs) constitute a family of proteins sharing a common structural organization characterized by an extracellular N-terminal end, seven hydrophobic alpha helices putatively constituting transmembrane domains and an intracellular C-terminal domain. GPCRs bind a wide variety of ligands that trigger intracellular signals through the activation of transducing G proteins (Caron, et al., Rec. Prog. Horm. Res. 48:277–290 (1993); Freedman, et al., Rec. Prog. Horm. Res. 51:319–353 (1996)).
More than 300 GPCRs have been cloned thus far and it is generally assumed that there exist well over 1,000 such receptors. Roughly 50–60% of all clinically relevant drugs act by modulating the functions of various GPCRs (Gudermann, et al., J. Mol. Med. 73:51–63 (1995)). Many of the clinically relevant receptors are located in the central nervous system.
Among the GPCRs that have been identified and cloned is a gene that encodes a protein homologous to the receptors of the DRR/RTA family. We called this receptor MAS receptor and described the structure of the gene as it exists in humans. However, the endogenous ligand for this family of receptors has not previously been identified (Cell: 45, 711–719 1986, JBC 273,11867–11873 1998, WO 99/32519).